The objectives of this research are to analyze the structure-function relationships of granulocyte-macrophage colony stimulating factor (GM-CSF) and to probe the molecular responses of cells in the GM series when they are stimulated by GM-CSF. This regulator is known to be involved in combating infections and even tumor resistance through its involvement in granulocyte-macrophage production and activation. Hence, it is essential to analyze the structural features of GM-CSF which confer its biological activity and specificity. Micro amino acid sequence analysis of GM-CSF will be undertaken using thin layer peptide mapping, radiolabeled GM-CSF and high pressure liquid chromatography for the analysis and identification of the PTH amino acid. Large scale production methods for the low molecular weight GM-CSF (23K) will be investigated using cell hybridization techniques. It is intended to produce sufficient GM-CSF to prepare fluorescent derivatives for the investigation of the processing of target cell receptors and to aid in the separation of GM-CFC. GM development is also modulated by other regulators. In particular there are molecules in post endotoxin serum, distinguishable from the GM-CSF, which promote GM differentiation in myeloid leukemic cells. These differentiation factors and GM-CSF from thigh muscle CM will be purified for direct comparison with GM-CSF from mouse lung CM. Myeloid cells will be fractionated to provide pure cell populations for GM-CSF binding and activation studies. Using a double laser fluorescent activated cell sorter GM-CFC, immature myeloid cells, granulocytes and monocytes will be purified to greater than 95% homogeneity. Hybridoma immunoglobulins specific for immature myeloid cells, and FITC-labeled GM-CSF will be used to separate GM progenitors from precursor cells in the other series. Mature normal and leukemic granulocytes will be used as a source of the GM-CSF receptor. Activation by GM-CSF of specific nuclear, cytoplasmic or membrane protein biosynthesis in normal and leukemic GM cells will be analyzed using two dimensional gel electrophoresis. Protein changes will be related to the functional activation, differentiation or proliferation status of the cells.